To produce a high-resolution image, a microscope requires a numerical aperture in excess of about 0.6. To form a sharp image of an object over its full lateral extent, the features of the object that are of interest must be within the depth of focus of the microscope. To try to maintain the full lateral extent of an important feature of an object within the depth of focus of a microscope, high-grade mechanical stages are typically used to achieve sufficient flatness and repeatability of travel. In a scanning microscope the problem of maintaining the feature within the depth of focus can be remedied in part by detection of the best focal plane for the feature and adjusting the position of the optical system along its optical axis relative to the stage to compensate for any changes in flatness as the feature is moved laterally with respect to the optical system. This solution can only be successfully implemented to the extent the stage can repeatably position the object along the optical axis. In any case, a numerical aperture of 0.6 or more can reduce the depth of focus to the sub-micrometer level, which challenges the capability of known mechanical stages.
Another problem encountered with high numerical aperture microscopes is that, since the numerical aperture of the illumination system should match the numerical aperture of the observation optics to maximize the optical resolution of the image, a high numerical aperture requires a relatively large space for the mechanical components of the stage. This is because of the need to accommodate the components of a high numerical aperture illumination system.
These problems become particularly acute in a recent innovation in microscopy known as a miniature microscope array (“MMA”) or, when applied to a common object, as an “array microscope”. In miniaturized microscope arrays, a plurality of imaging lens systems are provided having respective optical axes that are spaced apart from one another. Each imaging lens system images a respective portion of the object.
In an array microscope, a linear array of imaging elements is preferably provided for imaging across a first dimension of the object, and the object is translated past the fields of view of the individual imaging elements in the array, so that the array is caused to scan the object across a second dimension to image the entire object. The relatively large individual imaging elements of the imaging array are staggered in the direction of scanning so that their relatively small fields of view are contiguous over the first dimension. The provision of the linear detector arrays eliminates the requirement for mechanical scanning along the first dimension, providing a highly advantageous increase in imaging speed.
The MMA concept invites the corresponding concept of providing each imaging element with a corresponding trans-illumination element. For optimal effect, the numerical aperture of the illumination lens systems needs to be matched to the numerical aperture of their corresponding imaging elements. That is, if the illumination system transmits light to the object at angles greater than the acceptance angle of the imaging system, some of the light may be wasted, which reduces system efficiency. On the other hand, if the illumination system transmits light over a narrower angular range, that is, one that does not extend to the acceptance angle, the imaging system cannot take full advantage of its resolving power.
In a high numerical aperture array microscope it is desirable to pack the imaging elements of the array close together so as to acquire image data for contiguous parts of the object in the minimum scan time. However, a trans-illumination system places a limit on how close the corresponding illumination lens system can be packed and still provide the desired matching of numerical apertures. This is because the object must be supported by a slide or other transparent member that must be sufficiently thick to provide mechanical stability. Where the illumination system must project light through a glass substrate 1 to 1.5 mm thick, the working distance cannot be greater than that amount. To have a sufficiently long illumination system working distance, while maintaining the same numerical aperture as the imaging system, the diameter of the lens of the illumination system must be larger than the diameter of the lens of the imaging element. This means that providing each imaging element with its own illumination element requires either that sub optimal imaging element packing or sub optimal numerical aperture matching must be employed. However, in a related patent application Ser. No. 10/191,874, entitled SINGLE AXIS ILLUMINATION FOR MULTI-AXIS IMAGING SYSTEM, it has been disclosed that in a multi-axis imaging system such as an array microscope, a single axis trans-illumination system permits maximum packing of the imaging elements and optimum matching of the numerical aperture of the illumination system with the numerical aperture of the imaging elements, while providing a practical working distance for the illumination system. Thus, a single axis optical system may be provided for illumination, preferably having the same numerical aperture as the individual imaging elements and an exit pupil large enough to fill the collective contiguous fields of view of the imaging array.
Since an object of using an MMA ordinarily is to achieve a high-resolution image, the afore-mentioned problem of maintaining focus with a scanning, high numerical aperture microscope array is typically encountered. Also, due to the wide lateral dimensions of the array, a relatively large stage is required to accommodate the illumination system whether it is a single axis or multi-axis illumination system.
Accordingly, there is a need for a microscope stage that maintain high flatness and repeatability during lateral movement for scanning, and that provides room for relatively large trans-illumination optics.